Article Figures & Data
Figures
- Fig. 1.
[14C]serine (25 μM) accumulation by washed cells of P. gingivalis W50 (▴) and the serine transport mutant W50ST (♦) that were energized by tetrapeptide Gly-Gly-Tyr-Arg (100 μM) addition 5 min prior to radiolabeled serine addition. The effect of gramicidin addition (↓) on serine accumulation by P. gingivalis W50 is shown (▪). Serine accumulation by washed P. gingivalis W50 cells with no tetrapeptide addition is also shown (●). All experiments included 10 mM cysteine in the assay buffer. The data points represent the mean values from three separate experiments.
- Fig. 2.
Lineweaver-Burk double reciprocal plot of initial serine uptake rate versus initial serine concentration. Initial uptake rates were determined in washed P. gingivalis cells energized by addition of the tetrapeptide Gly-Gly-Tyr-Arg 5 min prior to [14C]serine addition at pH 7.5, 37°C. Data points represent the average of three to six determinations.
- Fig. 3.
Alignment of the deduced amino acid sequences of SstT from P. gingivalis (Pg) and E. coli(Ec). Shading indicates identical residues and bold indicates conservative substitutions. Conserved motifs found in the glutamate transport family (30) are boxed.
- Fig. 4.
Southern blot analysis of P. gingivalis genomic DNA. The DNA of the wild-type W50 (lane 1) and W50ST (lane 2) were digested with BamHI andPstI, and the resulting DNA fragments were subjected to agarose gel electrophoresis and Southern blot hybridization with atetQ (a) or sstT (b) probe.
Tables
- Table 1.
Concentrations of amino acids in P. gingivalis W50 continuous culture medium, 1.75% BHI broth containing 2% BSA and 4 mM (each) free serine, threonine, and arginine, before inoculation and in spent, cell-free medium
Residue Amino acid concentration (mM)a Before inoculation In spent medium Free form Peptide form Free form Peptide form Ser 5.1 11.3 0.1 5.4 Thr 5.1 12.2 0.0 6.8 Arg 5.3 7.5 0.1 3.6 Asx 1.1 19.9 0.7 10.1 Glx 1.6 33.4 1.3 16.8 His 0.3 5.9 0.2 3.1 Gly 0.5 7.7 0.6 3.5 Cys 3.1 12.1 2.9 8.2 Ala 1.4 21.9 1.2 13.8 Tyr 0.4 6.5 0.5 3.3 Val 1.2 13.4 1.6 6.5 Met 0.6 2.4 0.3 0.9 Trp 0.4 NDb 0.0 ND Phe 1.4 8.6 0.7 4.1 Ile 0.9 6.1 0.6 2.4 Leu 3.3 19.9 2.8 10.4 Lys 1.7 19.8 2.4 10.2 Pro 0.2 15.1 7.2 4.4 - Table 2.
Effects of EDTA, gramicidin, and sodium ion concentration on the rate of [14C]serine uptake by washed cells ofP. gingivalis
Buffer Sodium ion concentration (mM)a Rate of serine transport (%)b Pga 39 100 ± 16 Pga + EDTA 39 103 ± 2 Pga + gramicidin + EDTA 39 NDc PgaKd 0.35 6 ± 17 PgaNaKe 9.3 43 ± 2 Pga + NaCl 112 ND ↵a Determined by atomic emission spectroscopy.
↵b Rates are presented as a percentage of the control treatment. Experiments were conducted in triplicate, and the results are presented as the mean ± standard deviation.
↵c No uptake detected.
↵d PgaK buffer was essentially the same as Pga buffer, except that the buffering capacity was provided by KH2PO4 and the pH was adjusted with KOH.
↵e PgaNaK buffer was essentially the same as Pga buffer, except that the pH was adjusted with KOH.
